We plan to study the organization and expression of various mammalian polypeptide hormone gene systems: namely insulin; the glycoprotein hormones (chorionic gonadotropin, luteinizing hormone, follicle stimulating hormone, and thyroid stimulating hormone); and the growth hormone related proteins (growth hormone, chorionic somatomammotropin and prolactin). Both cDNA and genomic clones will be isolated from specific human and rat tissues using recombinant DNA methods and analyzed by nucleotide sequence analysis. The coding and noncoding regions (5' and 3' untranslated and intervening sequences) of related genes will be compared within and between species. The location(s) of the genes will be mapped by hybridization analysis of restriction endonuclease digests of cloned DNA isolated from bacteriophage lambda and cosmid gene libraries and also DNA from metaphase chromosomes separated with a fluorescence activated cell sorter. The cloned DNAs will be used as molecular hybridization probes to study the expression in cultured cells of the hormone genes in response to external stimuli, such as other hormones. Putative control regions in the cloned genes will be mutated in vitro by chemical, enzymatic, and/or oligonucleotide methods and introduced back into mammalian cells by cell transformation or by using SV40 as a vector. The expression of the "wild type" and various mutant genes in normally producing and non-producing recipient cells (e.g. growth hormone in GC versus HTC rat cells) will be measured using nucleic acid hybridization (for both DNA and RNA) and protein detection (polyacrylamide gel electrophoresis and immunological) methods. Other projects include studies on the structure and replication of the retrovirus genome and the isolation of the purine nucleoside phosphorylase gene and its expression in purine nucleoside phosphorylase deficient mouse teratocarcinoma cells.